The three main objectives of the proposed studies on myelin assembly are (1) characterization of the processing of galactolipids and phospholipids between synthesis and assembly into myelin, (2) characterization of the processing of proteolipid protein between synthesis and assembly into myelin, and (3) determination of effects of corticosteroid treatment on neural development, with emphasis on myelination. The possible role of Golgi membranes in metabolism of myelin galactolipids will be investigated by chase experiments in brain slices and by in vivo experiments with double isotope techniques. Processing of the choline and ethanolamine phospholipids will be examined by determining the subcellular distribution of the enzymes which synthesize the phospholipids, and of methyl transferases which convert ethanolamine to choline phospholipids. The enzymes which synthesize the lipids will be investigated with regard to membrane asymmetry by determining their susceptibility to proteolytic enzymes in intact microsomal vesicles. The distribution of total and newly synthesized galactolipids and phosphatidyl ethanolamine in mocrosomes from brainstem will be studied with galactose oxidase and TNBS, respectively, to determine what proportions of the lipids are exposed on the luminal vs. the cytoplasmic surface. These methods for localizing the lipids and the enzymes which synthesize them will be applied to fractions enriched in smooth endoplasmic reticulum and Golgi membranes. We will determine whether proteolipid protein is synthesized on free or membrane-bound polysomes, and apply the methods developed to identify the protein to studies similar in strategy to those used for the lipids. Previous observations that entry of proteolipid protein into myelin is more sensitive than basic protein to inhibition by puromycin and colchicine will be investigated by measuring synthesis of the proteins in brain slices and polysomes exposed to these agents. In continuing studies on myelination in coritcosteroid-treated animals, the timing and duration of hormone treatment will be varied. DNA content, myelin yield, and sulfatide synthesis at various ages will be followed as indicators of cell number and extent of myelination.